Past results from this ongoing research endeavor strongly support the theory that endogenous adenosine plays a significant physiological role to restrain the release of renin in response to a variety of stimuli. Two important questions regarding this theory, referred to as the adenosine brake hypothesis, linger: 1) Does the adenosine brake exist? and, 2) How does it operate? The first objective is to conduct a critical test of the adenosine brake hypothesis that should corroborate the in vivo existence of this regulatory system. Since adenosine inhibits renin release via A1 adenosine receptors, if endogenous adenosine functions to limit the renin release response to stimuli, then selective A1 receptor, but not A2 receptor, blockade should augment the renin release response to agents such as isoproterenol and PGI2. This prediction will be tested by using a series of novel A1 and A2 receptor antagonists that recently were evaluated and validated in this laboratory. The second objective is to investigate the mechanism of the adenosine brake on renin release. In this regard, the transmembrane negative feedback loop hypothesis will be tested as a potential explanation for how the adenosine brake on renin release operates. This hypothesis proposes that: 1) hormonal activation of adenylate cyclase (AC) stimulates renin release via cAMP-induced activation of protein kinase A; 2) stimulation of AC causes egress of cAMP out of juxtaglomerular cells and/or neighboring cells; 3) extracellular cAMP is degraded locally to AMP and hence to adenosine; 4) adenosine, via an A1 adenosine receptor, inhibits AC and consequently brakes the renin release response to the original hormonal stimulus. This hypothesis would be addressed in part by the same experiments used to test the existence of the adenosine brake, and will be further tested by determining: 1) whether A1 receptor blockade does not alter the renin release response to 8-bromo-cAMP; 2) whether PGI2 and isoproterenol increase renal cortical interstitial levels of cAMP, AMP, adenosine, inosine and hypoxanthine and whether these increases are appropriately altered by pharmacological agents; 3) whether cAMP and AMP inhibit renin release and whether this inhibition is appropriately modified by pharmacological agents; 4) whether infusions of cAMP and AMP appropriately alter renal cortical interstitial levels of purines and whether these changes are appropriately altered by pharmacological agents; and 5) whether inhibition of cAMP-dependent protein kinase attenuates the effects of isoproterenol and PGI2 on renin release and prevents A1 receptor antagonists from potentiating renin release responses to isoproterenol and PGI2.